S-alleles through PCR amplification of RNase and SFB regions with different primer combinations. The S-genotype of 66 apricot cultivars is reported, 41 for the first time. Forty-nine cultivars were considered self-compatible and 12 self-incompatible, which were allocated in their corresponding incompatibility groups. Additionally, the available information was reviewed and added to the new results obtained, resulting in a compilation of the pollination requirements of 235 apricot cultivars. This information will allow an efficient selection of parents in apricot breeding programs, the proper design of new orchards, and the identification and solution of production problems associated with a lack of fruit set in established orchards. The diversity at the S-locus observed in the cultivars developed in breeding programs indicates a possible genetic bottleneck due to the use of a reduced number of parents.In recent years, an important renewal of apricot cultivars is taking place worldwide, with the introduction of many new releases. Self-incompatible genotypes tolerant to the sharka disease caused by the plum pox virus (PPV), which can severely reduce fruit production and quality, are being used as parents in most breeding programs. As a result, the self-incompatibility trait present in most of those accessions can be transmitted to the offspring, leading to the release of new self-incompatible cultivars. This situation can considerably affect apricot management, since pollination requirements were traditionally not considered in this crop and information is lacking for many cultivars. Thus, the objective of this work was to determine the pollination requirements of a group of new apricot cultivars by molecular identification of the  Prunus armeniaca L.) belongs to the genus Prunus in the Rosaceae family. This crop was domesticated in different domestication events ca. 2000–3000 years ago in Central Asia, which is considered the center of origin of the crop , in which only one S-allele could be previously identified . Th. ThS-genRc-(F/R) ,77. PCR a et al. .S6/S9 and S1/S7, showed similar fragment sizes, specific primers based on the second intron of the RNase were used to distinguish between them. For the identification of the S6- and S9-alleles, the PruC2/PruC4R primer combination designed from P. avium S-RNase-cDNA sequences [S1 and S7, respectively [w/v) agarose gels and the DNA bands were visualized using the nucleic acid stain SYBR Green . For the identification of the S1-allele, the specific primers SHLM1 and SHLM2 were used following the protocol described by Herrera et al. [Taq DNA polymerase . Primers SHLM3 and SHLM4 were used for S7-allele identification. PCR reactions were performed with Phusion® High-Fidelity DNA Polymerase  according to Herrera et al. [Because two pairs of alleles, equences  was usedectively . PCR reaectively , but witectively . The ampa et al.  for Taq a et al. .cS- and S8-alleles using the primers SRc-(F/R) provides a fragment of similar size [SFB gene, were used to distinguish between both alleles [Since PCR amplification of the lar size ,56, the  alleles . The PCR alleles .cS- and S8-alleles. Self-(in)compatibility was established by laboratory-controlled pollinations and the observation of pollen tube growth under fluorescence microscopy [In cultivars ‘Samourai’ and ‘Water’, no amplification was produced with the primers AprFBC8-F/AprFBC8-R. Thus, controlled pollinations were carried out in these two cultivars to differentiate the croscopy . Self-pocroscopy , was usev/v) during 24 h, and conserved at 4 °C in 75% ethanol. After hand pollinations, pollen viability was evaluated. Pollen from each pollen donor was scattered on a solidified pollen germination medium [Flowers from each cultivar were collected at the balloon stage one day before anthesis and emasculated to avoid self-pollination. Pistils were placed on wet florist foam and maintained at laboratory temperature. After 24 h, a group of 20–25 flowers were hand-pollinated with the help of a paintbrush for each self- and cross-pollination . Pollen n medium  and poll2 during 10 min in sodium sulphite to soften the tissues [v/v) aniline blue in 0.1N K3PO4 [For histochemical preparations, the fixed pistils were washed three times for 1 h with distilled water and left in 5% sodium sulphite in distillated water at 4 °C for 24 h. Then, they were autoclaved at 1 kg/cm tissues . Pistils1N K3PO4  to obserPollen tube behavior was observed in at least 10 pistils in each self-pollination. Cultivars were considered as self-incompatible when pollen tube growth was arrested along the style in most self-pollinated pistils. On the other hand, when the pollen tube reached the base of the style in most self-pollinations, the cultivars were considered as self-compatible.S-allele genetic diversity, the S-RNase genotypes of the 60 cultivars identified herein were compiled with those of the 175 cultivars previously reported , allelic richness (Ar), and private alleles (Pa) were calculated for all countries on the traditional and cultivars from breeding programs using the adegenet 2.1.3 [S-allele was calculated in each country within each group and the results were plotted as a heatmap with the R package PopGenReport version 3.0.4 [S-alleles and their distribution by country, a contingency table of absolute frequencies of alleles by country was created and a chi-square test was performed with the “chisq.test” function using the R package “stats” v. 4.1.2. Due to the low number of observations in some countries, a Monte Carlo simulation with 2000 replicates was indicated.The number of alleles (Net 2.1.3 , and Popet 2.1.3  packageson 3.0.4 . To analS-alleles, in addition to the determination of the pollination requirements of the cultivars, can elucidate missing gaps in the evolution, domestication, and dissemination of the crop. The diversity at the S-locus observed in the cultivars developed in breeding programs indicates a possible genetic bottleneck due to the use of a reduced number of parents in breeding programs.Results reveal that a significant proportion of new apricot releases are self-incompatible and, therefore, require cross-pollination to produce fruit. Knowing the incompatibility relationships between cultivars will help breeders to select suitable parental genotypes in crosses. This information, combined with the flowering dates in each geographical area, will allow the selection of appropriate inter-compatible pollinizers for self-incompatible cultivars in the design of new orchards. The identification of the


Background: The role of preventive measures increases significantly in the absence of effective specific COVID-19 treatment. Mass population immunization and the achievement of collective immunity are of particular importance. The future development of public attitudes towards SARS-CoV-2 immunization depends significantly on medical students, as future physicians. Therefore, it seemed relevant to determine the percentage of COVID-19-vaccinated medical students and to identify the factors significantly affecting this indicator. Methods: A total of 2890 medical students from years one to six, studying at nine leading Russian medical universities, participated in an anonymous sociological survey. The study was performed in accordance with the STROBE guidelines. Results: It was found that the percentage of vaccinated Russian medical students at the beginning of the academic year 2021 was 58.8 ± 7.69%, which did not significantly differ from the vaccination coverage of the general population in the corresponding regions (54.19 ± 4.83%). Student vaccination rate was largely determined by the region-specific epidemiological situation. The level of student vaccination coverage did not depend on the gender or student residence (in a family or in a university dormitory). The group of senior students had a higher number of COVID-19 vaccine completers than the group of junior students. The lack of reliable information about COVID-19 vaccines had a pronounced negative impact on the SARS-CoV-2 immunization process. Significant information sources influencing student attitudes toward vaccination included medical professionals, medical universities, academic conferences, and manuscripts, which at that time provided the least information. Conclusion: The obtained results make it possible to develop recommendations to promote SARS-CoV-2 immunoprophylaxis among students and the general population and to increase collective immunity. The long-term pandemic of the new coronavirus infection caused by the single-stranded RNA-containing virus SARS-CoV-2 has become a serious challenge not only for the healthcare systems but also for the economies of all countries . UnpreceOver the years of the pandemic, a significant amount of scientific information has been accumulated on the SARS-CoV-2 pathogenesis, the virus biology, and its interaction with the human immune system, making it possible to create effective COVID-19 vaccines ,6,7. In A special risk group in the pandemic situation is medical students who have a high infection risk due to their frequent visits to different patients and emergency practice in coronavirus hospitals ,27,28. MA total of 2890 1st- to 6th-year medical students from nine leading Russian universities located in the regions with different epidemiological SARS-CoV-2 situation were included in a cross-sectional online anonymous survey conducted in the period of late September−early October 2021 . By thisThe study was carried out on the Google Forms platform  in accordance with STROBE guidelines. The reference to the questionnaire was distributed among students through social networks and Internet information channels. Using G*Power software statistical package   and baseRespondent sample representativeness in each of the nine universities is presented in https://xn--80aesfpebagmfblc0a.xn--p1ai/information/, accessed on 15 November 2021) were used to identify the correlation between student behavioral attitudes toward SARS-CoV-2 immunization in relation to morbidity, mortality, lethality, and vaccination population coverage of the corresponding Russian region.The up-to-date data provided by Russian Federal Service on Customers’ Rights Protection and Human Well-Being Surveillance . Minimum incidence and mortality were observed in the Tatarstan Republic (p < 0.05), where Kazan State Medical University is located (Moscow and Saint Petersburg led steadily with respect to numbers of patients and number of deaths per 1000 people among the Russian regions studied ( located .p < 0.05). The minimum value of this indicator was observed in the Kirov region (p < 0.05)  .COVID-19 vaccination coverage of the population of each region at the time of the study is shown in The original questionnaire was designed using prior scientific studies on vaccine and behavioral attitudes toward immunization, with new factors added ,33,34,35The work fully complied with the requirements of the local ethical committee of Sechenov University (Protocol No. 04-19 dated 6 March 2019) and the norms of the Declaration of Helsinki. All students were recruited on a volunteer basis and gave informed consent before the study. Respondents understood the survey purpose and were told how to fill in the questionnaire. No reward was offered to participants.p < 0.05. The results were counted twice by 2 independent researchers. The inter- and intragroup correlation agreement rates were greater than 95%; for this reason, all results were considered as the mean between 2 attempts by 2 researchers.Statistical data analysis was performed using SPSS 20.0  and MedCalc 11.5.00 . The minimal subject number needed for this study was calculated by means of power analysis. Descriptive statistics had a place for data from scales where the mean, median, standard deviation, and percentiles were calculated. Intergroup qualitative indicators were compared using Pearson’s χ-squared test or Fisher’s exact test. The normality of distributions was checked with Kolmogorov–Smirnov test. Quantitative differences were determined using the independent t-test for normal distribution or the non-parametric Mann–Whitney U-test when variables were non-normally distributed. For multiple-comparison, non-parametric Kruskal–Wallis test and parametric ANOVA were used. Pearson’s coefficient and Spearman’s rank test were calculated for correlation analysis. Statistics were considered significant at The mean respondent age was 20.99 ± 2.28 years (range of 16 to 37 years) . The surMost students consider COVID-19 a dangerous disease (89.4%) and express profound concern about the current unfavorable sanitary–epidemiological situation (89.9%) associated with the rapid spread of new coronavirus variants. Three-quarters of the respondents have already encountered this disease in their relatives (75.9%), and every third had experienced the death of a loved one (36.98 ± 5.79%). Almost half of the young surveyed people (48.6%) had experienced RT-PCR test-confirmed COVID-19 themselves, with one in two (46.6%) being diagnosed with moderate and severe form of this disease (according to computed tomography criteria). Most students (85.4%) are responsible with respect to following non-specific SARS-CoV-2 preventive measures, which primarily include wearing masks in public places, treating hands with antiseptic, and social distancing. Respondents rated the effectiveness of these anti-COVID measures at 4.73 ± 0.51 points .Most future specialists (65.0%) consider vaccination to be the most effective method for infectious disease prevention; every second respondent (57.3%) sees the immunization expediency even directly during a pandemic. However, only a little more than a third of all students (39.0%) expressed a definite positive attitude towards COVID-19 vaccination, and the same number of respondents (42.7%) have not yet fully decided on this issue, so only half of all respondents (52.5%) recommend their relatives, friends, and acquaintances to vaccinate against coronavirus infection.p > 0.05). Many future doctors (66.5%) believe that COVID-19 vaccine prophylaxis acts as a necessary guarantee of protection against moderate and severe disease forms, fatal outcomes, and post-infectious complications. Only a tenth of the respondents (10.9%) suppose that active immunization can prevent disease incidence.The effectiveness of SARS-CoV-2 specific prophylaxis was estimated by medical students at 4.87 ± 0.50 points , which did not differ significantly from the effectiveness of non-specific SARS-CoV-2 preventive measures  indicate that the COVID-19 vaccination need is primarily for at-risk individuals and adults. A quarter of respondents note that both adults and children should be vaccinated. One in six students (14.7%) is convinced that SARS-CoV-2 immunization is not indicated for anyone at all . Almost The highest percentage of (41.9%) medical students express the opinion that all COVID-19 vaccines are effective regardless of the origin country, 18.3% respondents are convinced of the good quality of only Russian vaccines, 24.3% respondents trust only imported vaccines. Every sixth medical student (15.5%) believes that all immunobiological agents are ineffective.Every third medical school student (31.9%) reported a clear lack of reliable information about anti-COVID vaccines, their composition, action mechanisms and contraindications. Sources of COVID-19 preventive vaccines, used by medical students for subsequent informed consent or refusal to participate in a vaccination campaign against COVID-19, were identified .Thus, medical students received most information about active SARS-CoV-2 immunization drugs from the Internet and the mass media. Half of the respondents, in order to make their choice (52.3%), relied on information obtained from medical professionals. The least importance in obtaining knowledge about vaccines was found for medical universities, scientific conferences and manuscripts, the role of which is reported only by a third of students, which is comparable to the number of students whose source of information is familiar. Half of future doctors (47.7%) are not satisfied with the quality of information they received at all.p < 0.05). The distribution of the vaccinated by the region in Russia is shown in p < 0.05).The COVID-19 vaccination coverage among Russian medical students was 58.8%, which did not significantly differ from the percentage of citizens who were vaccinated in the studied regions, which was 54.2% (p > 0.05), and was significantly higher in the group of senior students compared to junior students . It was found that in a significantly larger percentage of cases, students were motivated to receive the COVID-19 vaccination certificate than to prevent the disease and its severe course .The percentage of COVID-19-vaccinated students was not dependent on gender  and the single-component vaccine Sputnik-Light, based on viral vectors and developed by the N.F. Gamaleya Federal Research Center for Epidemiology & Microbiology . In secoThe most common side effects after COVID-19 vaccination were weakness (71.4%), injection site soreness (66.2%), fever (64.5%), headache (54.1%) and muscle aches (52.2%), which, as a rule, disappeared on their own within 2–3 days. In 14.8% of respondents, side effects were completely absent.The predominant reasons for not vaccinating were lack of awareness about COVID-19 vaccines and doubts about their efficacy and safety (15.3%), fear of side effects and post-vaccination complications (23.5%), presence of post-COVID immunity (15.3%) and medical withdrawal from vaccination due to existing contraindications (12.0%).Multicomponent correlation analysis made it possible to identify the factors affecting the COVID-19 vaccination coverage among Russian medical students. Thus, it was shown that the student attitude towards COVID-19 vaccination was almost independent of their opinion about the danger of this coronavirus infection (r = −0.009) and their concern about the current sanitary–epidemiological situation (r = −0.235). It was found that students were motivated to carry out specific prophylaxis not so much by an increase in the overall incidence in the region (r = 0.388) as by the incidence among students themselves (r = 0.573) and their relatives (r = 0.626). Increased mortality rates because of SARS-CoV-2 (r = 0.655) and, especially, the percentage of deaths among close people (r = 0.778) had a great effect. It should be noted that there is a strong positive correlation between the percentage of vaccinated medical students and the lethality rate because of SARS-CoV-2 in this region (r = 0.701). There was no relationship between the SARS-CoV-2 immunization coverage of the region population and the proportion of SARS-CoV-2 vaccinated medical students (r = −0.119).A pronounced negative impact on the SARS-CoV-2 immunization process was provided by the lack of information about COVID-19 vaccines (r = −0.638) and the unreliability of these data (r = −0.693), which determine the doubt about the vaccination effectiveness and safety (r = 0.674), and the fear of side effects and post-vaccination complications (r = 0.686). It was found that the percentage of students who decided to be vaccinated against SARS-CoV-2 was positively correlated with the percentage of students whose main information sources were medical professionals (r = 0.484), medical university (r = 0.584), and scientific conferences and manuscripts (r = 0.317). Information from the Internet (r = 0.164) and from familiar (r = −0.096) had no significant effect on student SARS-CoV-2 immunization attitudes. Students who received information mainly from the mass media vaccinated in a significantly lower percentage (r = −0.623). It should be emphasized that the percentage of young people who were negative about SARS-CoV-2 immunization (r = −0.450) and not fully decided on this issue (r = −0.722) was negatively correlated with the percentage of COVID-19-vaccinated students.It was found that the student COVID-19 vaccination coverage did not depend on their student residence, i.e., whether they lived in a family (r = 0.055) or in a university dormitory (r = −0.043). A negative correlation was noted between the percentage of COVID-19-vaccinated students and the proportion of students who exclusively prefer foreign vaccines (r = −0.322). The percentage of future specialists who recommended COVID-19 vaccination to everyone around them and who, accordingly, largely affect the formation of collective population immunity, was significantly correlated with the percentage of students who were positive about SARS-CoV-2 immunization and had been vaccinated themselves (r = 0.903).Study design and research results are summed up in flow chart diagram .According to recent studies, the understanding of the importance of the COVID-19 vaccine among medical students all over the world is very high, since COVID is a potentially severe acute respiratory infection ,37,38,39Hesitancy in accepting COVID-19 vaccination has a place among medical students in different countries and in some studies its prevalence even reaches 23.0–46.0% ,28,40. ITherefore, the real COVID-19 vaccination rate among Russian medical students in autumn 2021 was shown to be 58.8 ± 7.69%, which did not differ from the COVID-19 vaccination coverage among the general population at that time, and was significantly lower than the target value for collective immunity, which, according to actual data of the Russian Federal Service on Customers’ Rights Protection and Human Well-Being Surveillance was 80%. The number of COVID-19-vaccinated medical students worldwide was also not sufficient and according to systematic analysis of 40 original articles published between January 2020 and December 2021 averaged 61.9% .It was found that the main reason for COVID-19 vaccination hesitancy among medical students and the low vaccination rate in Russia is the lack of reliable information on COVID-19 vaccines, their composition, action mechanisms and contraindications, which determine the doubt about the effectiveness and safety of COVID-19 vaccination, and the fear of side effects and post-vaccination complications, which has also been emphasized by other scientists ,33,34. IFor the first time, it was found that the student COVID-19 vaccination rate was largely determined by the actual epidemiological situation of the region, and primarily by indicators such as mortality (r = 0.655) and lethality (r = 0.701), which may be related to medical students’ awareness of COVID-19 treatment options and care quality in corresponding regions. It was proved that student COVID-19 vaccination coverage rates were independent of gender and student residence. For the first time, our study demonstrated that the group of senior students had a higher number of COVID-19 vaccine completers than the group of junior students, which may be due to their greater involvement in coronavirus hospitals and more frequent contact with health care workers. Other authors also established that willingness to be vaccinated against COVID-19 among medical students generally increased with age and education .It has been shown that the percentage of students willing to recommend COVID-19 vaccination to the people around them and thereby contribute to increasing the collective immunity level is significantly dependent on the percentage of students who have been COVID-19 vaccinated. Many scientists consider that medical students are among the front-line medical professionals who meet patients. In addition, it is very important to achieve high rates of COVID-19 vaccination coverage in this group of health care providers, as they will recommend COVID-19 vaccination and counsel vaccine-hesitant people . Public One of our study limitations was the lack of result separation by faculty, as there is a perception that medical student knowledge varies between different specializations. We consider all respondents to have a high level of education corresponding to their course. In addition, the relatively small number of males compared to females may likely be a limiting factor on the study results. In future studies we will choose proportional numbers of students from all medical universities. Another limitation is that the survey was conducted under the condition that only Russian vaccines were included. For this reason, this study and its results must be understood as the initial stage of multi-central research for COVID-19 vaccination in order to understand its problems.The real COVID-19 vaccination rate among Russian medical students in autumn 2021 was 58.8 ± 7.69%, which did not differ from the COVID-19 vaccination coverage of the general population at that time, and was significantly lower than the target value for collective immunity. The main reason for low COVID-19 vaccination among medical students’ rate is the lack of reliable information about COVID-19 vaccines. Significant information sources influencing students’ attitudes toward vaccination include medical professionals, medical universities, academic conferences, and manuscripts. The group of senior students had a higher number of COVID-19 vaccine completers than the group of junior students.The obtained results support the following recommendations:(1) The primary task is to eliminate the information deficiency about COVID-19 vaccines among medical students using educational resources and, above all, the medical university, considering its significance in the forming adherence to SARS-CoV-2 immunologic prophylaxis. (2) SARS-CoV-2 immunologic prophylaxis education should be organized to all students, focusing on junior students. (3) Registration of foreign COVID-19 vaccines in Russia and development of official recommendations for their use will increase the percentage of vaccinated students and population. (4) A conscious decision by medical students to be vaccinated against SARS-CoV-2 will contribute to their competent explanation of immunologic prophylaxis essence to the population and increase collective immunity.


Soy protein isolate (SPI) is an attractive natural material for preparing wood adhesives that has found broad application. However, poor mechanical properties and unfavorable water resistance of wood composites with SPI adhesive bonds limit its more extensive utilization. The combination of lysine (Lys) with a small molecular structure as a curing agent for modified soy-based wood adhesive allows Lys to penetrate wood pores easily and can result in better mechanical strength of soy protein-based composites, leading to the formation of strong chemical bonds between the amino acid and wood interface. Scanning electron microscopy (SEM) results showed that the degree of penetration of the S/G/L-9% adhesive into the wood was significantly increased, the voids, such as ducts of wood at the bonding interface, were filled, and the interfacial bonding ability of the plywood was enhanced. Compared with the pure SPI adhesive, the corresponding wood breakage rate was boosted to 84%. The wet shear strength of the modified SPI adhesive was 0.64 MPa. When Lys and glycerol epoxy resin (GER) were added, the wet shear strength of plywood prepared by the S/G/L-9% adhesive reached 1.22 MPa, which increased by 29.8% compared with only GER (0.94 MPa). Furthermore, the resultant SPI adhesive displayed excellent thermostability. Water resistance of S/G/L-9% adhesive was further enhanced with respect to pure SPI and S/GER adhesives through curing with 9% Lys. In addition, this work provides a new and feasible strategy for the development and application of manufacturing low-cost, and renewable biobased adhesives with excellent mechanical properties, a promising alternative to traditional formaldehyde-free adhesives in the wood industry. The marrheating . Becauserheating .Soy protein isolate (SPI) is a by-product of the soybean oil industry . With thCurrently, chemical modification is one of the most viable modification strategies to enhance the performance of SPI adhesives . The cro2 group in lysine (Lys) reacts with the epoxy resin and therefore has high reactivity [The curing of epoxy resin requires selecting a suitable curing agent . Petroleactivity . Lys hasactivity . HoweverAs a result, the water-based glycerol epoxy resin (GER) was employed as a cross-linking agent to modify the SPI adhesive chemically. Amino acids were used as accelerators to enhance the interfacial bonding with the SPI adhesive. The combination of Lys with a small molecular structure as a curing agent for soy-based wood adhesives allows Lys to penetrate easily into wood pores and can result in better mechanical strength of soy-based composites. The effect of Lys as a curing agent cross-linking on the structure and performance of SPI and its composites was investigated. This knowledge could help to manufacture green biocomposites with low cost, good mechanical properties, and thermal stability using SPI and wood.−1 was related to the stretching vibration of O-H [2 and COOH, produced O-H groups that attested to the direct participation of this moiety in the cross-linking reaction. Thus, the intensity in the 3432 cm−1 peak increased, which can be seen from the variation between adhesives a and c–h [−1 belong to the C-H stretching bands of the saturated structural CH2 and CH3 groups, and at 1733 cm−1 were characteristic peaks corresponded to the C=O groups in COOH or COOR, mainly originating from amino acid small molecules [−1 for the C=O stretching (amide I), at 1525 cm−1 for the N-H bending (amide II), and at 1394 cm−1 for the superposition of C-N and N-H vibrations (amide III) [Fourier-transform infrared (FT-IR) spectra of SPI, GER, and soy-based wood adhesives are illustrated in n of O-H . When thn of O-H . In cont and c–h . The absolecules . The pea−1 was for the C-O-C, which mainly originated from GER and can be clearly seen in adhesive b. The reaction of GER dramatically reduced the intensity of the C-O-C absorption peak in the modified SPI adhesives c-h. [−1 were attributed to the characteristic peaks of epoxy group, which was the essential characteristic peak for the hydrophilic group of GER and the SPI in the cross-linking reaction [2 and -COOH groups of the protein molecule reacted sufficiently to form a cross-linking structure [The peak at 1070 cmves c-h. . The peareaction . Comparetructure .The XRD spectra of pure SPI and modified SPI adhesives are demonstrated in Comparing SPI and S/G-9%, new diffraction peaks were observed at 30.6° and 44.3° in S/G-9%, indicating that these two peaks belong to the added glycerol epoxy. The diffraction peaks were still observed at 31.5° in S/G/L-3%, and with the addition of Lys, the diffraction peaks belonging to GER could not be directly observed. The crystallinity of the SPI adhesives is shown in The apparent toughness tests of the pure SPI, adhesive modified with GER (S/G-9%), and adhesive modified with GER and Lys  are shown in 2, COOH, and -OH [Compared with the pure SPI adhesive, S/G/L-9% enhanced the apparent toughness significantly after adding the cross-linking agent. The hydrophilic groups on the SPI adhesive were weakened by the interaction between the epoxy group of GER and reactive groups, including -NH and -OH . The forSoy-based wood adhesives have the fluidity to wet and penetrate the wood . The appThe apparent viscosity decreased considerably with the addition of glycerol epoxy S/G-9% (135.5 Pa.s) compared to the pure SPI (276.7 Pa.s) adhesive, mainly being attributed to the addition of the low-molecular-weight glycerol epoxy acting as a lubricant between the molecular protein chains, resulting in a reduction in viscosity. Compared with the pure SPI adhesive, the viscosity of the modified adhesive (S/G/L-9%) was reduced by adding 9% lysine. This phenomenon was attributed to the lubricating effect of adding a small amount of amino acid, and too much amino acid affected the viscosity of protein adhesives.The effects of S/G-9%, S/G/L-3%, S/G/L-6%, S/G/L-9%, S/G/L-12%, and S/G/L-15% on the gluing performance of plywood were analyzed by investigating the penetration of adhesive into the substrate in the modified protein adhesives and the interfacial bonding ability of plywood .In The residual rate is a reflection of the water resistance of the adhesive . The resIn general, with the addition of the GER, the residual rate of the SPI adhesive was enhanced. This can be explained by the addition of GER consuming the hydrophilic groups in the adhesive system and developing a cross-linked network through the ring-opening cross-linking reaction, thus effectively improving the adhesive bonding strength . The resDry and wet shear strength tests were used to assess the influence of GER and Lys on SPI adhesives . With th2, COOH, and -OH. On the one hand, this chemical interaction decreased the content of hydrophilic groups in the adhesive. On the other hand, it formed a highly cross-linked network through the cross-linking reaction, which enhanced the adhesive bonding strength of the adhesive. When excessive amino acids were added, some amino acids did not participate in the reaction, which increased the adhesive’s solid content and destroyed the cross-linking between the GER and amino acids [The wet shear strength of the pure SPI adhesive was only 0.64 MPa, which did not meet the strength requirement of China Class II plywood (≥0.7 MPa) . After ano acids . The wetThe damage to the middle layer of plywood can reflect the adhesive bonding strength . The morThe wood destruction rates of plywood prepared by modified SPI adhesives were all superior to the plywood prepared by the pure SPI adhesives, which suggested that the addition of the GER and Lys enhanced the bonding strength of SPI adhesives. The most remarkable wood destruction rate was S/G/L-9%, which reached 84%. This phenomenon illustrated that the bonding strength of the plywood climbed and then declined with the addition of Lys under the condition of a fixed amount of the GER. Comparing S/G-9% (44%) and S/G/L-9% (84%), the destruction rate of S/G/L-9% was almost twice as high as that of S/G-9%. It was revealed that adding Lys into GER significantly enhanced the bonding strength of the plywood prepared by the modified SPI adhesive.The properties of modified SPI adhesives, including flexural strength and modulus were investigated . Among tThe DTG diagram shows that the adhesive degradation behavior comprised three reaction stages. After removing moisture in the first stage, the weak hydrogen and chemical bonds broke at 250–350 °C . The pep(Populus spp.) veneers  were purchased from ShunChuang Material Technology Co., Ltd. .The glycerin epoxy resin was obtained from Jiacheng Plastic Materials Co., Ltd. D-Lysine (98%) was purchased from Aladdin Chemical Reagent Co., Ltd. . SPI with a content of 90% was supplied by Chang Jing Sheng Wu Gongcheng Co., Ltd. . Poplar Firstly, 6.0 g SPI and 0.54 g GER were dispersed in 44 g DI water and mixed at 25 °C for 20 min. After adding Lys, the mixed solution was further stirred at 25 °C for 20 min to obtain the modified SPI adhesive. −1 with 32 scans. The adhesives were pre-ground into powder form, and the crystallinity was analyzed using the X-ray diffraction (XRD)  analysis, with the diffraction angle scan range from 5 to 60°. The microstructural morphology of the adhesive interface of the plywood was sprayed with gold and photographed with a scanning electron microscope (SEM)  under an accelerating voltage of 10 kV.Different adhesives were solidified in a blast oven at 120 ± 2 °C for the Fourier transform infrared spectroscopy (FT-IR)  analysis. The scanning range was 4000–400 cmCracks in the cured adhesives were measured to determine the toughness of the adhesives. Adhesive samples were uniformly coated on glass slides and cured for 2 h in an oven at 120 °C. The dried samples were recorded the appearance the cracks using a camera.2 atmosphere with a flow rate of 10 °C/min in the range of 30–600 °C.The thermogravimetric (TGA)  analysis characterized the thermal performance of SPI adhesives. The test was taken under NThe mechanical performance of plywood was conducted on a universal testing machine . The sample size was 100 × 25 mm (length × width), and the loading speed was 20 mm/min. The testing distance between the two fixtures was 50 mm, and the average value was obtained by examining 13 samples.2. The uncoated veneers were stacked between two veneers coated with adhesive according to the principle of vertical texture between adjacent veneers. Then, the adhesive-coated veneers were placed in a cold press after 10 min for prepressing. Subsequently, the plywood was pressed at 120 °C and 1.0 MPa for 5.5 min using a hot press . Then, the shear strength was assessed according to the Chinese national standard (GB/T 17657-2013) [Three-lay plywood was fabricated to assess the bonding performance of these adhesive compositions. The adhesive of various formulations was evenly coated on one side of the 1.5 mm–thick poplar veneer with a density of 200 g/m57-2013) . The dry−1 in increments of 10 s−1 at 25 °C was examined with a Brookfield rotational viscometer .The apparent viscosity of adhesives from 0 to 150 sM1 at constant weights, the samples were soaked in water at 60 ± 2 °C for 6 h, then dried at 105 ± 2 °C. Again, the constant weights were recorded as M2, and the residual rate was calculated as:The SPI adhesives were placed at 120 ± 2 °C, and the masses were recorded as The preparation of modified soy-based wood adhesives with Lys as a curing agent improved the cross-linking reaction activity and physical penetration of soy protein into the wood, thus effectively enhancing the interfacial adhesion between the soy protein adhesives and veneers. Furthermore, the Lys has the same amine group as petroleum-based curing agents, and is an eco-friendly alternative to agent epoxy resin curing in the SPI cross-linking agents. The modified SPI adhesives had enhanced mechanical properties, apparent toughness, and thermal stability. The modified SPI adhesive manifested that the pyrolysis temperature was increased, and the thermal stability was improved. Compared to the pure SPI adhesive, the residual weight of S/G/L-9% was 26.2 wt% at degradation temperatures up to 600 °C. Compared to the pure SPI adhesive, the dry/wet shear strength of plywood prepared by the S/G/L-9% adhesive was enhanced by 52.7% and 90.6%, respectively. The flexural strength and modulus of the plywood prepared by S/G/L-9% adhesive achieved 80.38 MPa and 3.91 GPa, respectively, superior to plywood prepared by SPI adhesives . Such Lys cross-linking greatly improved the dry/wet shear strength, thermal stability, and water resistance. Overall, this study avoids the environmental pollution caused by the extensive use of petroleum-based curing agents in adhesives. The effect of Lys cross-linking as a curing agent on the structure and properties of SPI and its composites was investigated, providing a theoretical basis for the preparation of new low-cost and ecological adhesives as an alternative to formaldehyde-based adhesives.


Aspergillus are known to possess galactomannan in their cell walls. Galactomannan is highly antigenic to humans and has been reported to be involved in the pathogenicity of pathogenic filamentous fungi, such as A. fumigatus, and in immune responses. In this study, we aimed to confirm the presence of D-galactofuranose-containing glycans and to clarify the biosynthesis of D-galactofuranose-containing glycans in Aspergillus oryzae, a yellow koji fungus. We found that the galactofuranose antigen is also present in A. oryzae. Deletion of ugmA, which encodes UDP-galactopyranose mutase in A. oryzae, suppressed mycelial elongation, suggesting that D-galactofuranose-containing glycans play an important role in cell wall integrity in A. oryzae. Proton nuclear magnetic resonance spectrometry revealed that the galactofuranose-containing sugar chain was deficient and that core mannan backbone structures were present in ΔugmA A. oryzae, indicating the presence of fungal-type galactomannan in the cell wall fraction of A. oryzae. The findings of this study provide new insights into the cell wall structure of A. oryzae, which is essential for the production of fermented foods in Japan.Filamentous fungi belonging to the genus  GM functions as a component of the cell wall in filamentous fungi  and fungal-type GM (FTGM) (via the glycosylphosphatidylinositol (GPI) anchor as a carrier molecule  is a polysaccharide composed of D-mannose (Man) and D-galactofuranose  . OMGM isM (FTGM) . FTGM haM (FTGM) , and itsM (FTGM) . FTGM ismolecule . By consmolecule .f is highly antigenic to humans and is commonly detected in the blood of patients with invasive aspergillosis  was analyzed in different strains of A. fumigatus, and the loss of Galf antigens was found to have little effect on pathogenicity -Galf was added to the N-glycan outer chain structures  spectrometry revealed the presence of an FTGM-like structure in A. oryzae.Thus, this study aimed to clarify the structure of Gal2.2.1.Aspergillus strains used in this study are listed in 4·7H2O, and 0.152% w/v KH2PO4, plus Hunter’s trace elements (pH 6.5). To cultivate A. oryzae NSPlD1  using A. oryzae RIB40 genomic DNA as the template and the primer pairs ugmA-1/ugmA-2 and ugmA-3/ugmA-4, respectively (A. nidulans pyrG (AnpyrG) marker was amplified by recombinant PCR using pHSG396-AnpyrG  to yield pPTR-II-ugmA.2.4.A. oryzae was prepared as previously described . After neutralization with 50% acetic acid, the samples were dialyzed overnight against distilled water. The purified samples were then lyophilized, resuspended in distilled water, and clarified using 0.45-μm-pore filters. The resultant samples were prepared as GM fractions.Total GM  from escribed . Briefly2.5.1H-NMR were exchanged twice in D2O with intervening lyophilization and then dissolved in D2O (99.97% atom 2H). The 1H-NMR spectra were recorded using a JNM-LA600 spectrometer  at 45°C. The proton chemical shifts were referenced relative to internal acetone at δ 2.225.Samples for 2.6.Aspergillus enzyme immunoassay  was used at a dilution of 1:10 to detect β-Galf.Immunoblotting was performed as previously described . The EB-2.7.Hyphal surface adhesion assay was performed as previously described with slight modifications . Briefly3.3.1.f-containing sugar chains in A. oryzae, Aspergillus luchuensis (Aspergillus awamori var. kawachi) and A. luchuensis mut. Kawachii (A. kawachii), galactomannoproteins were extracted from mycelia and subjected to immunoblotting to detect Galf-containing glycoproteins using the anti-Galf antibody EB-A2  as a query sequence. AO090012000855 of A. oryzae RIB40, RIB2604_01707610 of A. luchuensis RIB2604, and AKAW2_10730A of A. kawachii IFO4308 exhibited strong homology to A. nidulans UgmA .To investigate whether the genes encoding UDP-galactopyranose mutase are present in the genome of A. nidulans UgmA as a query sequence. The BLAST search identified the UDP-galactopyranose mutase of A. oryzae, which was named AoUgmA by A. oryzae. However, as AoUgmB encodes only 168 amino acids, it probably does not function as a UDP-galactopyranose mutase because its protein size is too small. Therefore, AoUgmB was excluded from the present analysis. Phylogenetic analysis revealed that UgmA is widely distributed in the subphylum Pezizomycotina and the phylum Basidiomycota. In the phylum Ascomycota, Saccharomycotina and Taphrinomycotina do not carry UgmA. These results are consistent with previously reported findings and with the fact that galactofuranosyl chains are not present in Saccharomycotina and Taphrinomycotina  and observed the colonies , a chitin-binding reagent -β-fGal--β-Gal and β-Galf--β-fGal--β-Galf structures, respectively, according to previous studies on A. fumigatus  and UgmA in the cytosol (glfB/ugtA homolog gene (AO090012000853), encoding a Golgi-localized UDP-Galf transporter, is conserved adjacent to ugmA in the A. oryzae genome, as reported for A. fumigatus  is present in the genome as a gfsA/B/C homolog encoding β--galactofuranosyltransferase -galactofuran side chain of FTGM in A. oryzae. AO090009000688 is conserved as a homolog of the Golgi-localized GDP-Man transporter gene, gmtA (cmsA (AO090010000463) and cmsB (AO090120000333), encoding α--mannosyltransferase, and anpA (AO090023000901), encoding α--mannosyltransferase, which are involved in the biosynthesis of the FTGM core mannan backbone, are conserved in the A. oryzae genome  have not yet been identified in filamentous fungi, it can be inferred that the aforementioned genes are essential for the biosynthesis of FTGM in A. oryzae. Recent studies have postulated that FTGM is transported to the cell surface via the GPI-anchor and then cross-linked to β-glucan by Dfg proteins, which are mannosidases belonging to the GH76 family -glucan, α--glucan, chitin, and galactosaminogalactan  from the Japan Society for the Promotion of Science  and by Grant-in-Aid for Early-Career Scientists from Japan Society for the Promotion of Science .The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Background and Objectives: Polysomnography is manually scored by sleep experts. However, manual scoring is a time-consuming and labor-intensive task. The goal of this study was to verify the accuracy of automated sleep-stage scoring based on a deep learning algorithm compared to manual sleep-stage scoring. Materials and Methods: A total of 602 polysomnography datasets from subjects  aged 19 to 65 years  were included in the study. The performance of the proposed model was evaluated based on kappa value and bootstrapped point-estimate of median percent agreement with a 95% bootstrap confidence interval and R = 1000. The proposed model was trained using 482 datasets and validated using 48 datasets. For testing, 72 datasets were selected randomly. Results: The proposed model exhibited good concordance rates with manual scoring for stages W (94%), N1 (83.9%), N2 (89%), N3 (92%), and R (93%). The average kappa value was 0.84. For the bootstrap method, high overall agreement between the automated deep learning algorithm and manual scoring was observed in stages W (98%), N1 (94%), N2 (92%), N3 (99%), and R (98%) and total (96%). Conclusions: Automated sleep-stage scoring using the proposed model may be a reliable method for sleep-stage classification. Sleep accounts for one-third of a typical human lifetime and plays a very important role in maintaining good health. It is widely known that adequate sleep is associated with mental function and health, including the improvement of attention, learning, cognition, and memory ,2. AdditA sleep test or polysomnography is generally recommended for identifying sleep status and detecting sleep diseases, such as sleep-related breathing disorders, parasomnia, and sleep-related movement disorders . PolysomAttended overnight full-channel ≥7), in-laboratory polysomnography (level 1) is the most accurate and reliable type of sleep test. However, it has disadvantages, such as sleeping overnight in a laboratory, high cost, “first-night” effects, and inconvenience , in-labo,9.To overcome these drawbacks, various automated or computer-aided sleep staging algorithms have been developed and tested ,16,17,18n = 397) of this dataset consists of subjects diagnosed with obstructive sleep apnea , and mean AHI was 22.9 ± 26.0 events / hour of total sleep time. The percentage of total sleep time in each sleep stage W, N1, N2, N3, and R was 16.5 ± 13.7, 15.2 ± 11.2, 50.2 ± 13.5, 2.7 ± 4.7, and 15.4 ± 6.3, respectively. The datasets were collected using polysomnography at a tertiary university hospital between November 2016 and January 2019. Data were excluded from the dataset when data analysis was not possible, such as when the data were flawed or missing.The present study was designed as a retrospective analysis of polysomnographic data collected by the Soonchunhyang University Bucheon Hospital. This study was approved by the Institutional Review Board of Soonchunhyang University Bucheon Hospital, and informed consent was waived (SCHBC 2018-12-007). All methods of our study were performed in accordance with the relevant guidelines and regulations, including the Declaration of Helsinki. We included 602 datasets from subjects  aged 19 to 65 years . About two-thirds  scoring manual (Version 2.3) . Six-chaTime- and frequency-domain features, namely line length, kurtosis, and spectrograms of EEG signals, were extracted from 30 s epoch bases. Frequency-domain features were defined as delta-band power spectrum density (PSD), theta-band PSD, alpha-band PSD, and beta-band PSD using a multi-taper spectral analysis method with 29 sub-epochs with lengths of 2 s and overlaps of 1 s to capture the influences of sleep spindle and K-complex signals . All PSDThe kappa value was used as a performance evaluation metric for our proposed model. Kappa is a statistical index to use to measure inter-rater reliability for categorical items . BecauseIn the machine learning field, the bootstrapping method is widely used to estimate the performance of machine learning algorithms. This method provides summary statistics, such as means and standard deviations, to algorithm developers. To obtain these statistics, a resampling technique is applied by sampling a dataset with replacement. The main advantage of this method is that the performance of an estimator can be presented with CIs. This feature is not available with other methods. The basic principle of this method is that inferences regarding a population can be estimated by resampling sample data . With anThe proposed model, named StageNet , can be divided into six modules. The overall system architecture is presented in Cohen’s kappa statistics were used to quantity the level of agreement between sleep expert scores and the outputs of StageNet based on Soonchunhyang University Bucheon Hospital data .https://www.rproject.org/, accessed on 20 March 2020).We used 1000 bootstrapped samples to calculate the median percent agreement with a 95% CI. All analyses were conducted using the R software  were used to evaluate the performance of the proposed algorithm. We used 482 datasets for training, 48 datasets for validation, and 72 randomly selected datasets for testing.A confusion matrix for the proposed model is presented in The estimation results of the bootstrapping method are presented in High overall agreement between the proposed model and manual scoring can be observed for stages W (98%), N1 (94%), N2 (92%), N3 (99%), and R (98%). The average overall agreement between the two scoring methods for these sleep stages is 96%.The results of our experiments revealed good concordance rates and high overall agreement between manual sleep-stage scoring and automated sleep-stage scoring based on StageNet.n = 72) [n = 40). In this study, we also used the kappa value as an evaluation index to verify the performance of a newly developed automated sleep-stage scoring algorithm and observed a kappa value of 0.85. Compared to earlier validation studies on automated sleep-stage scoring, the results of this study revealed excellent outcomes. Stepnowsky et al. [n = 20). Zhang et al. [n = 5213). They calculated a kappa value of 0.82.Even though a standard for sleep-stage scoring was provided by the AASM, the scoring results of sleep technicians may differ based on ambiguity in brain waves and human errors. Danker-Hopfe et al.  investign = 72) . Deng etn = 72)  assessedy et al.  analyzedy et al.  evaluateg et al.  estimateVarious classifiers based on automated or computer-aided sleep-staging algorithms have been developed  ,16,17,18Most validation studies on automated sleep-stage scoring have reported that N1 is the hardest sleep stage to classify among the five sleep stages . The results of this study also demonstrate that the concordance rate for stage N1 is lower than those for stages W (94%), N2 (89%), N3 (92%), and R (93%). There are several possible reasons for these results. First, after scoring stage N2, a scorer continues to score the following epochs with low-amplitude, mixed-frequency (LAMF) EEG signals without sleep spindles or K-complexes according to the AASM scoring manual . TherefoThis validation study has a few limitations. First, it was designed as a retrospective evaluation of an automated sleep-stage-scoring system. Although this study was conducted using a relatively large dataset compared to other studies, additional polysomnographic data should be accumulated to increase the accuracy of deep learning algorithms for sleep-stage scoring. Second, this study was conducted with subjects aged 19 to 65 years. Future studies on subjects younger than 19 years of age or over 65 years of age are needed. Additionally, our clinical trial was conducted using subjects from a single institute. Therefore, further evaluation of patients at other institutes is required.A newly developed automated deep learning algorithm called StageNet exhibited excellent performance and may be a reliable tool for sleep-stage scoring in adults aged 19 to 65 years. However, additional research is required to improve the accuracy of the deep learning algorithm for sleep-stage scoring.


The need of palliative care at the end-of-life in the emergency departments (ED) is growing. The study aims to investigate the epidemiology of patients who died during care in ED using nationwide database, and to estimate the need for palliative care in the ED. A retrospective observational study was conducted using the National Emergency Department Information System (NEDIS) database. Patients who died during ED care between 2016 and 2019 were included. Palliative care-eligible disease was defined as cancer (C00–C99 of ICD-10), chronic respiratory disease , chronic liver disease , and heart failure . Among the 36,538,486 ED visits during 4 years, 34,086 ED deaths were included. The crude incidence rate of ED deaths per 100,000 person-year was steady between 16.6 in 2016 and 16.3 in 2019 (p-for-trend = 0.067). Only 3370 (9.9%) ED deaths were injury, while 30,716 (90.1%) deaths were related to diseases. The most common ED diagnosis was cardiac arrest (22.1%), followed by pneumonia (8.6%) and myocardial infarction (4.7%). In cases of disease-related ED deaths, about 34.0% stayed longer than 8 h in the ED (median (interquartile range): 4.5 (1.9–11.7) h) and 44.2% received cardiopulmonary resuscitation (CPR) at end-of-life time. A quarter of the disease-related ED deaths were diagnosed with palliative care eligible disease: cancer (16.9%), CLD (3.8%), HF (3.5%), and CRD (1.4%). Cancer patients received less CPR (23.4%) and stayed longer in the ED (median (interquartile range): 7.3 (3.2–15.9) h). Over the past 4 years, more than 30,000 patients, including 5200 cancer patients, died during care in the ED. A quarter of disease-related ED death were patients with palliative care-eligible condition and more than 30% of them stayed longer than 8 h in the ED before death. It is time to discuss about need of palliative care in the ED. This is, however, still in an introductory stage and is provided only in limited places and to limited subjects5. In Korea, the need for palliative and end-of-life care is increasing after the enactment of “Act on Hospice and Palliative Care and Decisions on Life-Sustaining Treatment for Patients at the End-of-Life” (hospice act) in 20186.The importance of palliative and end-of-life care is emphasized7. Some people die in ED from results of unexpected events or from sudden development of acute illnesses such as acute myocardial infarction or traumatic brain injury, while some patients with chronic diseases visit the ED to control rapidly worsening symptoms and serious complications of treatment and die in the ED9. Death during emergency care in the ED can be unavoidable for critically ill patients; however, there are little information about how many patients and who died in the ED10.The emergency department (ED) is a specialized space for treating acute medical problems and saving lives, and therefore mortality rate in the EDs is relatively higher than that of general wards13. People in proximity to death use more acute healthcare services than people who are not14. Among them, patients diagnosed with cancer and respiratory diseases are more likely to use acute-based healthcare service such as ED15. More than half of elderly patients who passed away in the ED had serious chronic illnesses and required palliative care17. Many ED visits in these terminally ill patients are considered avoidable, therefore, the need for palliative care at the end-of-life in ED is emphasized in many countries19. In the Unites States, there is a guideline for hospice care-eligible patients who visit the ED, and ED-initiated palliative care is recommended for patients with advanced cancer to improve the patients’ quality of life21. United Kingdom also reported end-of-life care recommendations for dying patients in the ED22. However, there is currently insufficient information on how patients who need end-of-life care are being treated in the ED and how many patients with incurable diseases received life-sustaining treatment such as cardiopulmonary resuscitation (CPR) in the ED before death. To improve the palliative care at end-of-life for terminally ill patients, it is necessary to understand the epidemiologic characteristics of patients who died in the ED23.As prevalence of the advanced disease increases, there is an increase in ED visits at end-of-life for patients with terminal cancer or chronic illnessesThe purpose of this study is to investigate the epidemiologic characteristics of patients who died in the ED using representative nationwide database and to identify the reasons of ED visits of those patients. The study intends to estimate the need for palliative care at end-of-life in the ED and to accumulate the baseline data for future research.26. The information includes patient demographics, prehospital, and ED treatment. All patient-related information was automatically transferred from each ED to a central government server within 14 days of discharge from the ED or hospital. The trained coordinators designated in each institution managed the data uploading process. The NEDIS data is updated by the National Emergency Medical Center and is approved annually by National Statistics for data quality management.This is a retrospective observational study using data from the National Emergency Department Information System (NEDIS) database. The Ministry of Health and Welfare constructed this nationwide ED-based database in 2013. NEDIS collected administrative and clinical information of all patients who visited EDs in real time from a total of 402 EDs across the country in 201927.Korea has a total population of 50 million, while the annual death rate was about 0.3 million in 2019. More than 70% of deaths occurred in medical institutions, and 14% at homes. The most common causes of death were cancer, heart disease, and pneumonia28.The National Health Insurance (NHI) of Korea covers the entire population for inpatient, outpatient, and ED treatment. EDs are classified into 3 levels according to resources and capacity , which are defined by the Ministry of Health and Welfare: 38 Level 1 EDs, 125 Level 2 EDs, and 239 Level 3 EDs , that are operational as of 2020. Level 1 ED provide the highest level of emergency care services in the country, and is designed to accommodate and to provide definite care to critically ill and severe emergency patients. Level 1 and 2 are emergency centers which take charge of treating critical ill patients, and Level 3 is in charge of general urgent care and primary responses 24 h a day, 7 days a week. Under this healthcare system, EDs are open to all beneficiaries without restriction, and patients have high accessibility to the ED care29. The hospice act defined four disease groups as target disease for palliative care: cancer, Acquired Immunodeficiency Syndrome (AIDS), chronic respiratory disease, and chronic liver cirrhosis as target disease for palliative care6. However, it is still in the introductory stage; there are not many providers of hospice ward and home-based hospice. The consultation-based hospice is mainly for inpatients and outpatients, and rarely applied to patients in ED.Hospice and palliative care was first introduced to Korea in 2015 with inpatient hospice beds. Consultation-based hospice care for non-hospice wards and outpatients started as a pilot project in 2017 and home-based hospice care was officially started in 2020The study subject were patients who died during care in the ED  in Korea for 4 years from January 2016 to December 2019. Death on arrival (DOA), patients who had already had cardiac arrest at ED arrival and did not receive any resuscitation efforts, patients who visited ED for non-medical reasons such as issuance of medical certificates, and patients who had unknown information of age or reason for ED visit were excluded.Based on the reason for ED visit, all ED deaths were initially classified into a disease-related group and injury-related (injury) group. For disease-related ED deaths group, the specific reasons for ED visits were classified based on the ED diagnosis codes of the international classification of diseases, 10th edition (ICD-10) in the ED discharge records written by emergency physicians.30. Palliative care-eligible disease was defined based on the ED diagnosis: cancer , chronic respiratory disease , chronic liver disease (K70–K77), and heart failure (I50).Palliative care-eligible disease was defined based on the hospice act and literature review: cancer, chronic respiratory disease, chronic liver disease, and heart failure32.The following variables were collected from the NEDIS database to investigate the characteristics of ED deaths: 1. patient demographics , 2. prehospital and ED information , route of ED visit , region of ED , and level of ED ), 3. care and outcomes , life-sustaining treatment , ED diagnosis , ED disposition , and ED death)A descriptive analysis was conducted to investigate epidemiologic characteristics of the study population. Categorical variables were presented as counts and proportions and differences across groups were tested by Chi-square test. Continuous variables were presented as medians and interquartile ranges (IQR) and differences across groups were tested by Wilcoxon rank-sum test.The characteristics were compared according to the reason for ED visit (disease-related and injury-related). The most common diagnoses were investigated for the disease-related ED deaths group and cancer-related ED deaths.33.The epidemiologic trends of all ED deaths and ED deaths of patients with palliative care-eligible conditions were investigated. The crude and the age- and sex- standardized incidence rates per 100,000 person-years were calculated. Direct standardization method was used using the 2020 mid-year census population as the standard populationNEDIS database collected during 2016–2019 were analyzed in July 2021. All statistical analyses were conducted using SAS software version 9.4 . Statistical significance was taken as P < 0.05.This study was approved by the Institutional Review Boards of Seoul National University Hospital , and the requirement for informed consent was waived due to the retrospective nature of this study. Patient information was anonymized prior to analysis. All methods were performed in accordance with relevant guidelines and regulations.The National Emergency Medical Center under the Ministry of Health and Welfare was involved in the design and conduct of this research, but it was not possible to involve patients in our research.Between 2016 and 2019, the total number of ED visits was 36,538,486 of which total death toll was 117,256 (0.3%). Among them, 34,086 ED deaths were included as a study population, excluding cases that were already in cardiac arrest at the ED entrance  and cases with unknown information on age (n = 3) and reason for ED visit (n = 1945)  died from injury while 30,716 (90%) died from diseases. The median age of ED deaths was 75 (63–83) years old. 32.2%  had an alert mental status upon ED arrival. Median (IQR) ED-LOS was 4.4 (2.0–11.2) h and 32.8%  stayed in the ED over 8 h before death. CPR was provided in 45.4% and endotracheal intubation was performed in 48.6%. Compared to the injury-related ED deaths group, the disease-related ED deaths group was older (76 (64–83) vs. 68 (54–79) years old, p < 0.01) and had longer ED-LOS (4.5 (1.9–11.7) vs. 3.6 (2.0–6.9) h, p < 0.01) , followed by pneumonia (8.6%) and myocardial infarction (4.7%). In cancer patients, the most common diagnoses excluding cancer diagnoses (42.7%) were pneumonia (6.5%), followed by cardiac arrest (6.3%) and dyspnea (3.8%)  were ED deaths of patients with palliative care-eligible: 16.9% (n = 5187) was related with cancer, 1.4% (n = 416) with chronic respiratory disease, 3.8% (n = 1176) with chronic liver disease, and 3.5% (n = 1076) with heart failure. The ED deaths of patients with palliative care-eligible conditions had longer median ED-LOS, compared to ED deaths of patients without palliative care-eligible conditions (p < 0.01). Regarding life-sustaining treatment, 2327/7855 (29.6%) received CPR and 2707/7855 (34.5%) received endotracheal intubation before death. Cancer patients received relatively less life-sustaining treatment: 23.4% received CPR and 27.1% received endotracheal intubation  of the 30,716 disease-related ED deaths did not receive CPR prior to death. Patients who did not receive CPR had longer ED-LOS than those who received CPR (6.5 (2.6–16.0) vs. 3.0 (1.6–6.8) h, p < 0.01). Of the ED deaths that received CPR, 17.2% were diagnosed with palliative care-eligible diseases . . The age- and sex- standardized incidence rate of ED deaths per 100,000 person-years decreased from 19.7 in 2016 to 17.1 in 2019 (p-for-trend < 0.001). Both disease (17.5 to 15.6) and injury (2.2 to 1.4) were on the decline (p-for-trend < 0.001)  before death, compared to other patients. Cancer-related ED deaths accounted for 16% of disease-related ED deaths: half of them died after staying for more than 8 h in the ED and a quarter of them received CPR before death. This study found that nearly 8500 patients die in the ED every year and the annual incidence rate of ED deaths was about 17 per 100,000 person-year in Korea. To the best of current knowledge, it is the first study to investigate the magnitude of ED deaths nationwide.27. Since the ED is for providing acute care for critically ill patients, death in the ED is unavoidable. However, not much is known about patients who die in an ED34. In this study, only 10% of the ED deaths were caused by injuries such as traffic accidents or falls. This was smaller than a single center study done in Switzerland10. 90% of the ED deaths were disease-related patients, and one quarter of them had diagnosis codes subject to palliative care. Previous studies have reported that 30 ~ 50% of death in the ED were related to serious chronic illnesses or palliative care needs35. Since this study classified only four representative diseases as palliative care-eligible diseases, size of the palliative care subjects may be smaller than other studies. Theoretically, all patients with imminent death are eligible for palliative care, including those who die in the ED2.In Korea, the death rates per 100,000 person-years in 2019 were 574.8 for all cause of death and 158.2 due to cancer, accounting for 27.5% of all deaths32. Other studies reported that, among ED visits, discharged patients stayed for median 2.5 h and hospitalized patients stayed for 4 h37. Patients who die in ED may stay for relatively longer periods of time in the ED and utilize more resources than patients who survived. Among all ED deaths, about half of ED deaths received CPR (45.4%) and endotracheal intubation (48.6%) before death in this study. It aligns with other studies in that more than 50% of the patients died in the ED without CPR40. Moreover, 30% of ED deaths with palliative care-eligible diseases and a quarter of cancer-related ED deaths received CPR before death, which is 4–7 times higher than that of a German study41. CPR is a life-saving treatment for acute stage patients, but it could be a life-sustaining treatment with little meaning for incurable and terminally ill patients. A hospital-level study or qualitative study is needed to know how life-sustaining treatment is provided in individual patients. The results of this study show the possibility that there still is a substantial unmet need for palliative care in the ED and the need for follow-up studies43.In this study, ED deaths happened after a median 4.4 h in the ED, and 30% of disease-related ED deaths occurred after 8 h in the ED. It was previously reported in US that all ED visit patients stayed for median 4 h in the ED, and that more than 8 h of stay is defined as a long stayIn this study, about half of ED deaths with palliative care-eligible diseases and cancer-related ED deaths stayed over 8 h before death in the ED. During ED care, 30% of ED deaths with palliative care-eligible diseases and a quarter of cancer-related ED deaths received CPR before death. More attention would be needed on how to provide end-of-life care in times of expected death in the ED. This discussion encompasses not only which care should be provided for dying patients, but also whether it would be suitable for such patients to stay in ED for such long times. This study showed overall epidemiologic characteristics of ED deaths using a nationally representative emergency database. Considering that patients who died within 72 h after visiting the ED, as well as ED death, are also eligible for palliative care, the need for palliative care at the end-of-life in the ED would increase . Further study is needed to determine whether patients with palliative-care eligible conditions have a plan before visiting the ED and can receive proper end-of-life care through this. As a limitation of the studies using a large database, it is necessary to comprehensively interpret the results of this study by referring to the results of studies focusing on individual patients. Second, whether the patient had a disease subject to palliative care was determined based on the diagnosis codes in the ED medical records. Whether that disease was previously diagnosed or first diagnosed in the ED is unknown. Third, the NEDIS data does not collect information of test results and medication prescription. It is difficult to know whether the dying patient received sufficient supportive care in the ED before death, such as pain control. Fourth, Korea has the NHI service is implemented, and palliative care services are in the introduction stage. It is difficult to generalize the result to other countries with different healthcare system and accessibility to palliative care.In conclusion, more than 30,000 people died during care in the ED over the past four years. At least a quarter of disease-related ED deaths were owed to palliative care-eligible diseases. More than 30% of disease-related ED deaths and 40% of cancer-related ED deaths happened at least 8 h after arrival in the ED. About half of disease-related ED deaths and 30% of ED deaths with palliative care-eligible diseases received CPR before death. Increased efforts to understand death in the ED may improve end-of-life care in the ED.Supplementary Tables.


At 22°C, spch-5 had lower transpiration and warmer leaves than the wild type. However, at 30°C, this mutant developed larger stomata and thinner leaves, paralleled by a notable cooling capacity, similar to that of the wild type. Despite their low stomatal density (SD), spch-5 plants grown at 30°C showed no photosynthesis or growth penalties. The behavior of spch-5 at supra-optimal temperature exemplifies how the effect of very low stomatal numbers can be counteracted by a combination of larger stomata and thinner leaves. Furthermore, it provides a novel strategy for coping with high growth temperatures.The impact of global warming on transpiration and photosynthesis would compromise plant fitness, impacting on crop yields and ecosystem functioning. In this frame, we explored the performance of a set of Arabidopsis mutants carrying partial or total loss-of-function alleles of stomatal development genes and displaying distinct stomatal abundances. Using microscopy and non-invasive imaging techniques on this genotype collection, we examined anatomical leaf and stomatal traits, plant growth and development, and physiological performance at optimal (22°C) and supra-optimal (30°C) temperatures. All genotypes showed thermomorphogenetic responses but no signs of heat stress. Data analysis singled out an extremely low stomatal abundance mutant,  Arabidopsis thaliana) seedlings, PHYTOCHROME-INTERACTING FACTOR 4 (PIF4) acts as a central hub where other signaling pathways involving light, hormones, and circadian clocks, as well as epigenetic processes related to chromatin remodeling converge to modulate the morphogenetic responses to supra-optimal temperature  and thinner leaves, contribute to promote cooling, what plants do mostly through transpiration . Transpiexchange . When stcenarios . The modn flecks . This wogsmax) depends on the number, size, and spatial distribution of the stomata on the leaf surfaces  and actual (gs) stomatal conductance with physiological processes, such as photosynthesis and transpiration  and supra-optimal (30°C) growth temperatures, a set of Arabidopsis mutants carrying hypomorphic or severe alleles for stomatal development genes, which display distinct stomatal abundance phenotypes. We examined (i) anatomical traits, such as SD, stomata and stomatal pore size, and leaf thickness and structure; (ii) plant growth by measuring bolting time, leaf numbers, the projected rosette area and dry weight (DW) in the various genotypes grown under the two conditions; (iii) physiological performance of the mutants in the two conditions through non-invasive imaging techniques, including their photosynthetic activity and their leaf temperature, as an estimation of transpiration. All genotypes showed the thermomorphogenetic responses described for Arabidopsis, albeit to different extents, and no signs of heat stress. Through this analysis, we singled out an extremely low stomatal abundance mutant previously isolated in our laboratory, A. thaliana (L.) Heynh., accession Columbia-0 (Col-0), epf1-1  to avoid potential selection bias by temperature during seedling establishment. For short-term supra-optimal treatments, the plants were grown under optimal conditions for 15 dag and moved to 30/24°C day/night for the last 3 days prior to measurements. All the plants were cultivated in growth chambers, on pots containing soil Compo Sana , and watered three times a week.The seeds were germinated under optimal conditions, as follows: 22/18°C day/night temperature, a 16-h photoperiod, 120 μE m0) and after (Fm) a 1-s saturating light pulse of 2,000 μE m–2 s–1. Subsequently, the plants were light adapted to 450 μE m–2 s–1 light for 10 min. During this light adaptation, saturating light pulses were provided at set times in order to record Ft and Fm’ before and after each light pulse, respectively. PSII maximum efficiency  and effective quantum efficiency of PSII  and non-photochemical quenching (NPQ) of absorbed light in PSII  during the light adaptation and at steady state were calculated according to The activity of the electron transport chain of the chloroplast was analyzed by chlorophyll fluorescence kinetics using a customized kinetic imaging fluorometer Open FluorCam FC 800-O , as previously described . BrieflyThermal images of the whole plants were recorded using a FLIR A305sc camera  vertically positioned 30 cm above the plants, according to Reflectance spectra in the 400- to 1,000-nm range of plant rosettes were collected using a Pika L hyperspectral imaging camera . The obtained spectra were used to calculate vegetation indexes (VIs) and other reflectance parameters on fully developed leaves. The parameters taken in consideration are related to different aspects of plant fitness, leaf composition, and structure  and are bolting), which was determined at 23 dag for plants grown at 22°C, and at 18 dag for those grown at 30°C. The number of fully developed leaves was registered for each plant. For the determination of the projected plant area, the plants were scanned in a standard office scanner and the images were analyzed with ImageJ 1.49v . DW of whole rosettes was measured after drying in an oven at 60°C for 2 days.Plant growth was analyzed in terms of morphology and plant biomass. Measurements were performed at 15 dag, with the exception of DW at bolting  microscopy in fixed fully expanded organs , as described in –2) was calculated in the fixed organs by counting stomata in two areas of 0.4 mm2 located on both sides of the median axis of the cotyledon or leaf, modified from n = 7). Stomatal size as previously defined  was measured as the length of the pore at the longest axis (μm). The maximum stomatal pore area (amax) was defined as a circle with the diameter equal to p (amax = π p2/4) and expressed in μm2. Stomatal pore depth (l) was estimated by single guard cell width. All measurements were made in micrographs of fixed organs using ImageJ  as the potential area available for gas exchange with the following equation , D is the SD value determined for each leaf, while l  and amax were the genotype average values. For each plant, the adaxial and abaxial gsmax were calculated and used to obtain the total for the leaf .where 2 were cut with a razor blade and immediately fixed in 1% paraformaldehide, 2% glutaraldehide, and 50-mM cacodilate (pH = 7.4) for 30 min under a vacuum. The samples were post-fixed in 1% OsO4 with 1% K3Fe(CN)6 for 1 h. The samples were dehydrated in subsequent EtOH solutions . The samples were finally embedded in EMbed 812 . Semithin 0.5-μm transections were obtained with ultramicrotome Reichert Jung S  and stained in 1% toluidine blue O and 1% borax. Sections were observed under an inverted microscope Leica DMi6000 (Leica Microsystems) and images analyzed by Adobe Photoshop . The average leaf thickness was calculated from leaf volume to area ratio (LVA) measured on micrographs as the volume corresponding to a 200-μm wide region of a 1-μm thick section, according to Leaf thickness was measured on micrographs of fully expanded leaf transections. Leaf samples of approximately 4 mmt-test was used for comparing two means. For multiple comparisons, the data were analyzed by one-way ANOVA followed by either the Tukey’s or Dunnett’s T3 post hoc test, or the Kruskal–Wallis test followed by the Dunn’s test. Principal components analysis (PCA) and hierarchical clustering on principal components (HCPC) were carried out on normalized and standardized variables using RStudio version 1.0.136 . The parameters considered for PCA and subsequent HCPC were selected following the criteria of no redundancy (to avoid overfeeding the PCA) and maximum-explained variance.Unless otherwise specified, statistical analyses were performed using the SPSS version 28.0 software . All experiments were carried out three times, providing comparable results. The data shown were from a representative experiment. Normality of the data and homogeneity of variance were verified by the Shapiro–Wilk and Levene tests, respectively. Student’s The symbols and descriptions of the parameters used in this study are listed in To evaluate the impact of SD on physiological traits under optimal growth temperature (22°C), we first selected eight mutant genotypes, displaying a range of SD values, using Col-0 as the wild-type control. In cotyledons, they encompass a broad SD variation, ranging from about 20–280% of the SD values in Col-0 . Some ofV/FM), PSII activity (by ΦPSII), and capacity for energy dissipation in the PSII (by NPQ)  . The orgion NPQ,  that, hoion NPQ, . A slightyledons . In contn as NPQ  and onlyof Col-0 . The sddtyledons  and tmm-n leaves . Remarkatyledons . NPQ meatyledons .spch mutants were significantly warmer. Deviating from this tendency, tmm-1 leaves showed the same temperature than those of Col-0. Consistent with the behavior of leaves, cotyledons of the spch mutants displayed the highest temperature values. The sdd1-3 and tmm-1 mutations, both causing high SD increases, led to much colder cotyledons than Col-0. A lower temperature was observed in epf2-3 cotyledons, while the rest of the mutants with moderate SD increases showed no significant differences with respect to Col-0.Leaf temperature mostly agreed with the SD values characteristic of the various genotypes, yet only some mutations impacted on cotyledon temperature . In geneer-116, with increased SD; tmm-1, with increased SD but presenting a large fraction of clustered stomata; plus spch-2 and spch-5, with SD decreased to a different extent.In order to assess the possible effects of SD alterations on plant growth and physiology, further experiments were carried out on a selection of genotypes in which the plants were grown at supra-optimal temperature (30°C). From the initial panel, genotypes with no or mild alterations in photosynthetic parameters were chosen: spch-5, which formed 6.2 leaves at optimal temperature and 8 at supra-optimal temperature. However, the rest of the genotypes produced a lower number of leaves than Col-0 or spch-5 when grown at 30°C, yet only er-116 had fewer leaves at 22°C.Growth was assessed for these genotypes in two temperature conditions (22 and 30°C) by measuring number of leaves, projected the rosette area and DW in the plants of 15 dag . As descspch-5, even though smaller than Col-0 as measured by the projected rosette area and DW when grown at 22 and at 30°C, developed the same DW  had produced about 400% more biomass than the same genotype grown at 30°C (at 18 dag). It is worth noticing that the nditions .spch-2 having leaves more elongated than the rest of genotypes and er-116, showing more compact rosettes and rounder leaves. In contrast, spch-5 and tmm-1 showed no changes in their morphology relative to the wild type.t-test p < 0.05), except for tmm-1. Compared to Col-0, the decreases were lower in spch mutants (14–15%), slightly larger in er-116 (21%), and marginal in tmm-1 (4%). In the adaxial side, only high stomatal abundance genotypes showed a decrease in SD at 30°C (t-test p < 0.05), being more pronounced in er-116. Although SD changes were more prominent in the abaxial side, the ratio of stomata on the abaxial and adaxial leaf surfaces  in all genotypes was similar in both growth conditions  or is even larger  than the amax of the high-SD mutant tmm-1 when grown at 30°C. In contrast, spch-2 had the smallest stomata and amax values in both epidermis and conditions, in spite of having a higher SD than spch-5 . Growth at supra-optimal temperature decreased the palisade layer in both genotypes but reduced the spongy tissue only in Col-0 (t-test p = 0.002), resulting in mesophylls with thicker spongy than palisade layers in Col-0  but of nearly identical thickness in spch-5 . It is worth noting that the palisade layer (the primary photosynthetic tissue) of spch-5 was, compared to Col-0, thicker at 22°C and equal at 30°C . Interestingly, while supra-optimal temperature also caused an increase in the thickness of the adaxial epidermis in both genotypes (t-test p < 0.00001), the abaxial epidermis became thinner only in spch-5 leaves (t-test p = 0.0003).As expected from previous findings , the leaof Col-0 , quantifof Col-0 . This anue types . While agsmax; see Section “Materials and methods”) can be calculated from stomatal anatomical traits. To estimate the theoretical potential for transpiration, we calculated gsmax for the adaxial and abaxial sides and the total gsmax(gsmaxT) at the two growth temperatures in fully expanded leaves of our genotype panel  stomatal conductance  had a strong positive contribution to PC1, whereas the gsmaxT and the ratio SDab/SDad, to a lesser extent, had a strong negative contribution to PC1. For PC2, DWbolting, leaf thickness, and the inhibition of PSII (FV/FM and NPQ), together with the VIs related to leaf structure (mRENVDI), plant vigor and stress , and pigments (PSSR2) had a strong positive contribution. The amax in the abaxial side of the leaf and leaf temperature , together with VIs related to pigment contents (CARI and ANTH2), had a strong negative contribution , and Cluster V consisted of Col-0 and the low SD spch mutants. Genotypes grown under 30°C clustered similarly with the exception of spch-2, which had its own cluster III, closer to high SD mutants instead of clustering with Col-0 and spch-5. It is remarkable that spch-5, with distinct anatomical differences as compared to Col-0  grouped together with the wild type, and both genotypes behaved very similarly when grown at supra-optimal temperature.The phenotypes shown by the five genotypes grown at optimal and supra-optimal temperature were analyzed by HCPC. The optimal number of clusters was found by cutting the generated tree, following the inertia gain criterion. Thus, genotypes were grouped in five clusters, represented in r = 0.7; epf1-1, epf2-3, er-116, and er-105) or severe (sdd1-3 and tmm-1), showed various alterations of PSII behavior  equal in spch-5 than in Col-0 in this leaf side, while the adaxial value was lower in the mutant (gsmaxT), we found that spch-5 overall values were lower, although its potential abaxial conductance was significantly higher than in Col-0, and despite having the same abaxial:adaxial SD ratio. Thus, although at supra-optimal temperature, spch-5 stomata experienced significant anatomical changes in comparison to Col-0, they did not translate into a better theoretical cooling capacity. However, indirect determination of transpiration showed that, at supra-optimal temperature, the mutant holds values indistinguishable from the wild type, what was surprising because, at optimal temperature, spch-5 was a warm, low transpiration mutant. This unexpected finding might be explained by the decrease in leaf thickness elicited by growth at supra-optimal temperature, which was observed in all genotypes as previously reported for Col-0 . There is clear evidence for a developmental coordination between epidermal and inner leaf tissues  and, morspch-5 grouped with the wild type, spch-2 had its own cluster very close to that of the high-SD mutants. The main traits explaining the variance among genotypes were stomatal size , the activity rate of photosystem II (ΦPSII), theoretical maximal conductance (gsmaxT), and the SD ratio between abaxial and adaxial sides for the first dimension. In the second dimension, the main traits were biomass at bolting time (DWbolting), leaf thickness, and several traits related to inhibition of photosynthesis, plant vigor, and stress, as well as the abaxial maximal pore size (amax) and leaf temperature. The close clustering of spch-5 and Col-0, particularly at 30°C, was surprising, given their significant anatomical differences and the important contribution of anatomical traits to our PCA and cluster construction. It is worth noticing that growth at 30°C results in spch-2, reaching lower values than Col-0 in number of leaves, the projected rosette area, DW and DW at bolting (spch-2 is only similar to Col-0 in SD but shows lower stomatal size and lower amax in both epidermis. This concurs with a faster NPQ induction (spch-2 mutation, i.e., on leaf development could alter mesophyll anatomy and, therefore, water and CO2 conductance and photosynthesis (as reviewed by spch-5 and spch-2 for relevant physiological, growth, and anatomical parameters and for their response to supra-optimal temperature. Their molecular lesions affect separate SPCH domains, the DNA-binding domain in spch-5, and the C-terminal of yet uncharacterized function in spch-2 (The developmental and functional effects of growth at supra-optimal temperature are supported by the results of PCA with a selection of anatomical, physiological, and VIs traits, as the hierarchical clustering classified the phenotypes first according to the temperature of growth . Within  bolting . Regardi bolting , spch-2 nduction , particun spch-2 . How thespch-5 in comparison with Col-0 further indicates that characteristics essential for fitness at supra-optimal temperature, such as cooling capacity and biomass accumulation, can be achieved through diverse anatomical solutions. spch-5 exemplifies how genotype-specific differential thermo-morphogenetic responses convert a low SD, warm, and slow-developing mutant into a successful genotype that performed similar to a wild-type plant under non-favorable conditions.Our results show that the anatomical and morphological responses to supra-optimal are genotype specific and not simply a consequence of SD. The behavior of the very low-SD mutant The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.MP-B and MM designed the experiments and analyzed the data with contributions of CF and MB. MP-B performed physiological, growth, and leaf thickness analysis. JI-M, AM-F, and AM determined the stomatal phenotypes. MM, MP-B, and CF wrote the manuscript with inputs from MB. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.